Colorimetric determination of l‐asparaginase enzyme activity most commonly uses Nessler's reagent, but often yields unreliable results due to precipitation of tetraiodomercurate salts. Interference caused by some commonly used inorganic and organic compounds of biochemical buffers, reagents and microbiology fermentation media was determined. The stabilising effects of tartrate and polyvinyl alcohol upon the impact of these salts was next evaluated using the following two‐step assay in 96‐well microtitre plates: 160 μL of 50 mmol L⁻¹ Tris–HCl pH 8.6, 32 μL of 100 mmol L⁻¹ l‐asparagine, 16 μL analyte and 128 μL water were added to each well. After incubation at 37 °C for 1 h, 16 μL of 1.5 mol L⁻¹ trichloroacetic acid was added to stop the reaction. A 35 μL volume was next transferred to a fresh well containing 35 μL Nessler's reagent, 35 μL stabiliser solution (4 mmol L⁻¹ disodium tartrate and 10 mg L⁻¹ polyvinyl alcohol) and 245 μL water. After incubation at room temperature for 10 min, absorbance was recorded at 436 nm and the enzyme activity calculated by extrapolation of a (NH₄)₂SO₄ calibrated standard curve.

中文翻译：

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使用Nessler试剂进行简单，准确的比色法测定l-天冬酰胺酶活性的改进方法

比色法测定l-天冬酰胺酶的活性最常用的是Nessler试剂，但由于四碘草酸酯盐的沉淀，经常会产生不可靠的结果。确定了由生化缓冲液，试剂和微生物发酵培养基中一些常用的无机和有机化合物引起的干扰。接下来，在96孔微量滴定板中使用以下两步测定法评估了酒石酸盐和聚乙烯醇对这些盐的影响的稳定作用：160μL的50 mmol L ^-1 Tris-HCl pH 8.6，32μL的100 mmol向每个孔中添加L ^-1 l-天冬酰胺，16μL分析物和128μL水。在37°C下孵育1小时后，得到16μL的1.5 mol L ^-1加入三氯乙酸以终止反应。接下来将35μL体积转移到含有35μLNessler试剂，35μL稳定剂溶液（4 mmol L ^-1酒石酸二钠和10 mg L ^-1聚乙烯醇）和245μL水的新鲜孔中。在室温下孵育10分钟后，在436 nm处记录吸光度，并通过外推（NH ₄）₂ SO ₄校准的标准曲线来计算酶活性。